Biochip with biosensors and fluidic devices

ABSTRACT

A biochip includes a substrate, where the substrate includes at least one hole extending from a first surface of the substrate to a second surface of the substrate opposite the first surface, and where the substrate comprises a microfluidic channel pattern. The biochip further includes a surface modification layer over the substrate. Additionally, the biochip includes a sensing wafer bonded to the substrate, where the sensing wafer has one or more modified surface patterns having different surface properties from each other.

PRIORITY CLAIM

The present application is a continuation of U.S. application Ser. No.13/838,905, filed Mar. 15, 2013, the disclosure of which is incorporatedherein in its entirety.

FIELD

This disclosure relates to biochips. Particularly, this disclosurerelates to biochips having biosensors and fluidic devices.

BACKGROUND

Biosensors are devices for sensing and detecting biomolecules andoperate on the basis of electronic, electrochemical, optical, andmechanical detection principles. Biosensors that include transistors aresensors that electrically sense charges, photons, and mechanicalproperties of bio-entities or biomolecules. The detection ofbiomolecules can be performed by detecting the bio-entities or thebiomolecules themselves, or through interaction and reaction betweenspecified reactants and bio-entities/biomolecules. Such biosensors canbe manufactured using semiconductor fabrication processes, can quicklyconvert electric signals, and can be easily applied to integratedcircuits (ICs) and microelectromechanical systems (MEMS).

Biochips are essentially miniaturized laboratories that can performhundreds or thousands of simultaneous biochemical reactions. Biochipscan detect particular biomolecules, measure their properties, processsensed signals, and may even analyze the corresponding+data directly.Biochips enable researchers to quickly screen large numbers ofbiological analytes for a variety of purposes, from disease diagnosis todetection of bioterrorism agents. Advanced biochips use a number ofbiosensors along with fluidic channels to integrate reaction, sensingand sample management. While biochips are advantageous in many respects,challenges in their fabrication and/or operation arise, for example, dueto compatibility issues between the semiconductor fabrication processes,the biological applications, and restrictions and/or limits on thesemiconductor fabrication processes.

BRIEF DESCRIPTION OF THE DRAWINGS

Aspects of the present disclosure are best understood from the followingdetailed description when read with the accompanying figures. It isemphasized that, in accordance with the standard practice in theindustry, various features are not drawn to scale. In fact, thedimensions of the various features may be arbitrarily increased orreduced for clarity of discussion.

FIG. 1 is a cross-sectional view of a biochip in accordance with variousembodiments of the present disclosure.

FIGS. 2A-2B are flow charts of various embodiments of methods offabricating a biochip device according to one or more embodiments of thepresent disclosure.

FIGS. 3-5, 6A-6B, and 7 are cross-sectional views of partiallyfabricated biochip devices constructed according to one or more steps ofthe methods of FIG. 2A according to one or more embodiments of thepresent disclosure.

FIGS. 8-9, 10A-10I, and 11A-11G are cross-sectional views of partiallyfabricated biochip devices constructed according to one or more steps ofthe method of FIG. 2B according to one or more embodiments of thepresent disclosure.

DETAILED DESCRIPTION

It is to be understood that the following disclosure provides manydifferent embodiments, or examples, for implementing different featuresof the invention. Specific examples of components and arrangements aredescribed below to simplify the present disclosure. These are, ofcourse, merely examples and are not intended to be limiting. Moreover,the formation of a first feature over or on a second feature in thedescription that follows may include embodiments in which the first andsecond features are formed in direct contact, and may also includeembodiments in which additional features may be formed interposing thefirst and second features, such that the first and second features maynot be in direct contact. Further still, references to relative termssuch as “top”, “front”, “bottom”, and “back” are used to provide arelative relationship between elements and are not intended to imply anyabsolute direction. Various features may be arbitrarily drawn indifferent scales for simplicity and clarity.

Some biochips involves various bioreceptors which react with variousbiological material of interest in one or more wells. One approach is totag a reaction with a fluorescent or phosphorescent bioreceptor thatemits a detectible photon. A coordinated approach would encode thesensor to a location on the biochip, so that a positive reaction andphoto-detection would be correlated to the location to determine thenature of the reaction, for example, identity of the biologicalmaterial. The signal may be optical, magnetic, electrical or amass-sensitive measurement such as surface acoustic wave or microbalanceweights. A random approach is to encode the sensor with differentfluorescence, phosphorescence, or otherwise detectible anddifferentiable sensors. A positive detection would be correlated to thetype of signal transduced to determine the nature of the detection. Thesignal transduced may be photons, for example, a different wavelengthlight is generated for different biological materials or surface plasmonresonance.

More advanced biochips involve not only biosensors, but also variousfluidic channels to deliver biological material to the sensors. Thefluidic channels may be a part of a microfluidic system that includespumps, valves, and various measurement devices such as flow meters,pressure transducers, and temperature sensors. Because a biochipcombines electrical processing and fluid processing, fluid handlingability has to successfully integrate within a semiconductor chipenvironment. A potential use of biochips is as a lab-on-a-chip—wheremedical professionals can use a small biochip to perform testing in thefield, obtain results contemporaneously, and proceed with treatment orfurther analysis without retreating to a laboratory. Especially formedical professionals working in remote areas where sample preservationmay be difficult, lab-on-a-chip devices can save lots of time spenttraveling and waiting. These lab-on-a-chip devices are often single-use,or disposable, devices. As such, the manufacturing costs have to be lowto be economically viable.

Semiconductor processing often involves baking, curing, and exposingvarious surfaces to plasma energy and radiation energy. At hightemperatures (i.e., above about 75 degrees Celsius) and/or highenergies, these processes would damage or destroy bioreceptors andsurface modification layers, which usually are delicate bio-molecules orvery thin layers of surface chemistry. For example, the bioreceptors maybe antibodies/antigens, enzymes, nucleic acids/DNA, cellularstructures/cells, and biomimetic receptors. The surface modificationchemistry may include thin layers (one or two molecules thin) ofhexamethyldisilazane (HMDS), 3-aminopropyl triethoxysilane (APTES),agar, or hydrogel.

Thus, the bio-functionalization of surfaces when bio-molecules areattached, are often performed after all the semiconductor processes arecompleted. In some designs, the fluidic channels are formed directly onthe semiconductor substrate, usually silicon wafer, along with thebiosensors. In other designs, the fluidic channels are formed on afluidic substrate that is subsequently bonded to the sensing wafer withthe biosensors. The fluidic channel formation, usually etching a trenchor via into the substrate, can damage the biosensors, bioreceptors orsurface modification layer. When a high temperature bonding process isused, the bioreceptors are deposited on the interior walls of thefluidic channels after the bonding process and the fluidic channels areenclosed, usually by flowing through each biochip a high concentrationof bioreceptors through the fluidic channel surfaces having someaffinity for the bioreceptors. However, the density of bioreceptors ishard to control and the process is slow and wasteful of bioreceptors andreagents. In some cases, the bound-bioreceptor density varies throughoutthe biochip or a batch of biochips (not uniform) as the concentrationsin the reagents change. The process may be limited to a single type ofbioreceptors because there the flow surfaces cannot be easily patternedafter the substrates are bonded and thus multiple detections are limitedto the random approach discussed above. For these reasons, the processis costly and disposable units are not economically viable.

The various embodiments of the present disclosure contemplate awafer-level process and a biochip that allows surface chemistry and/orbio-functionalization to occur before the bonding of the fluidicsubstrate and the sensing wafer at about room temperature withoutdamaging the exposed surfaces with high temperature, high energysemiconductor fabrication processes. The use of two substrates—thefluidic substrate and the sensing wafer substrate-introduces flexibilityin the choice of bonding material because one of the substrates may bedesigned to hold the more delicate bio-materials and the other substratemay be exposed to semiconductor processes that may otherwise damage thedelicate bio-materials. The fluidic substrate includes microfluidicchannels and inlets/outlets. The fluidic substrate is formed separatelyfrom the sensing wafer and total cycle time is reduced. One or more ofthe various method embodiments of the present disclosure may beperformed in a semiconductor fabrication facility and are compatiblewith the complementary metal-oxide-semiconductor (CMOS) process.

FIG. 1 is a cross-section of a biochip 100 in accordance with variousembodiments of the present disclosure. The biochip 100 includes afluidic substrate 101 and a sensing wafer substrate 103 with a bondingmaterial 105 between. The bonding material 105 is a biologicallycompatible material that does not react with the analyte, bioreceptors,or any testing fluid. The bonding material may be a photoresist. In oneexample, the photoresist is polysilsesquioxane (PSQ). In anotherexample, the bonding material is 3-aminopropyl triethoxysilane (APTES).PSQ and APTES are commercially available materials, for example, fromSigma-Aldrich® of St. Louis, Mo.

The fluidic substrate 101 has a fluidic inlet 107 and a fluidic outlet109 through the fluidic substrate 101. The fluidic substrate 101 has afirst surface 121 and a second surface 123. While labeled as inlet andoutlet, the flow of 107 and 109 may be reversed. The first surface 121includes microfluidic channel patterns, shown as channels and wells 111,113, 115, 117, and 119, referred to herein collectively as cavities. Thevarious cavities 111, 113, 115, 117, and 119 are connected to each othervia various pathways and may be different sizes depending on the designof the biochip. Some of the cavities may match a biosensor, providing awell having a sensing surface. Some of the cavities may be a fluidicreservoir. Some of the cavities may include bioreceptors that wouldreact detectibly with an analyte. The features between the cavities onthe fluidic substrate 101 are island features, for example, islandfeatures 127, 129, 131, and 133. According to various embodiments, theisland features 127, 129, 131, and 133 include a flat portion that isbonded to the bonding material 105 and hence through the bondingmaterial 105 to the sensing wafer 103. In some embodiments, the islandfeatures 127, 129, 131, and 133 are a portion of the fluidic substrate101 and the microfluidic channels 111, 113, 115, 117, and 119 are formedby etching into the fluidic substrate 101. In other embodiments, theisland features 127, 129, 131, and 133 are formed on the fluidicsubstrate 101 by depositing and patterning a material or by selectivelydepositing a material.

The fluidic substrate also includes a first surface modification layer145. The first surface modification layer 145 conformally covers all ofthe first surface 121 of the fluidic substrate 101 or a pattern on thefirst surface 121. In FIG. 1, the first surface modification layer 145covers all surfaces inside a fluidic channel, for example, channels andwells 111, 113, 115, 117, and 119, with exception of any surface coveredby a second surface modification layer 147. The first surfacemodification layer 145, shown in FIG. 1, may optionally cover thesidewalls of the fluidic inlet and outlet 107/109. The first surfacemodification layer 145, shown using dotted lines, may also optionallycover a top of the island features 127, 129, 131, and 133 and biochipedges 149 and 151. The biochip edges 149 and 151 include scribe linesfrom which each die containing a biochip would be separated from eachother. The biochip edges 149 and 151 are sufficiently wide such that thebiochip remains hermetically sealed between the fluidic substrate 101and the sensing wafer substrate 103 after dicing.

The first surface modification layer 145 is a bio-compatible materialthat provides a surface property to facilitate the operation of thebiochip. The surface property may be chemical, mechanical, orelectrical. For example, the first surface modification layer 145 mayprovide a hydrophilic, a hydrophobic, or a surface chemistry such asaffinity for particular functional groups. According to variousembodiments, the first surface modification layer 145 may behexamethyldisilazane (HMDS), silicon oxide, amino-silane (for example,3-aminopropyl triethoxysilane (APTES)), certain metal oxides, agar,or—hydrogel (for example, polyethylene glycol (PEG) hydrogel).Patterning of the first surface modification layer 145 may be performedto exclude it at the top of the island features 127, 129, 131, and 133if the first surface modification layer 145 does not adhere to thebonding material 105. In the example of HMDS, the HMDS renders the firstsurface 121 hydrophobic. During operation of the biochip, water-basedanalyte does not adhere to the hydrophobic surface and is directed tothe sensing surfaces which may be coated with a hydrophilic material. Inother embodiments, a hydrophilic coating is used for another purpose.

The fluidic substrate surface 121 may also include a second surfacemodification layer 147 that provides a different surface property thanthe first surface modification layer 145. If used, the second surfacemodification layer 147 is a different material from the first surfacemodification layer 145 and may be hexamethyldisilazane (HMDS), siliconoxide, silane, 3-aminopropyl triethoxysilane (APTES), certain metals ormetal oxides, agar, or polyethylene glycol (PEG) hydrogel. In someembodiments, the second surface modification layer 147 may be patternedon an externally observable or electrically detectible location on thefirst surface 121. In some embodiments, the second surface modificationlayer 147 is used to bind bio-molecules having certain functional groupsto specific wells or channels. During biochip operation, the boundbio-molecules can participate in reactions with analyte where either thereaction or result of the reaction is detectible. In some examples, thesecond surface modification layer 147 may be a metal or compound thathas affinity for certain functional groups, i.e. gold or platinum forthiol groups.—Another example is the coating of Poly-L-Lysine on glassor plastic-ware for promotion of cell adhesion. The thicknesses of thefirst surface modification layer 145 and the second surface modificationlayer 147 may be different.

The bonding material 105 between the fluidic substrate 101 and thesensing wafer 103 may be any bio-compatible adhesive, glue, polymer,epoxy, bonder, or solder that can provide a hermetic seal betweenchannels and wells. The bonding material 105 is compatible with CMOSprocesses and may be patterned. The bonding material 105 allows bondingof the fluidic substrate 101 and the sensing wafer at low temperatures,less than about 150 degrees Celsius or less than about 100 degreesCelsius. In one example, the bonding material 105 ispoly(phenylmethyl)silsesquioxane (PSQ). PSQ can be patterned like aphotoresist using a photolithographic process or printed in a patternusing a jet printer. Once activated using oxygen-containing plasma, PSQcan bond to a silicon containing substrate at room temperature. Inanother example, the bonding material 105 is 3-aminopropyltriethoxysilane (APTES). APTES can be deposited in vapor phase orprinted. Once activated using a catalyst, APTES can covalently bond to asilicon-containing substrate at room temperature. Suitable catalystsinclude water, toluene, and phosphate buffered saline (PBS) solution.

The sensing wafer 103 includes a patterned sensing surface 139 and oneor more biosensors 125. The biosensors 125 may be a biologicalfield-effect transistor (BioFET), an optical sensor (for example, a CMOSsensor), electromagnetic biosensors, electrochemical biosensors, andmechanical sensors (for example, mass sensitive sensors or motionsensitive sensors). The biosensors 125 may sense electrical, optical, ormechanical properties on the sensing surface or in the analyte.Electrical property biosensors measure a voltage potential or electricfield. Optical property biosensors measure intensity and wavelength of alight radiation. Mechanical property sensors measure a differentelectrical or optical value based on difference in mass or motion.Examples include piezoelectric sensors, laser Doppler vibrometers,Hall-effect sensors, and capacitive micromachined ultrasonictransducers. The biosensor 125 is connected through an interconnectstructure 135 to one or more electrodes 137. The electrodes 137 providepower and input/output of electrical signals to the biochip. Theelectrodes 137 may be bumps such as a ball grid array, copper pillars,or solder material. In various embodiments, the electrodes 137 allows areading of the data collected from the sample and may further allowcontrol of the biological analysis. The biochip 100 may be inserted intoa module having corresponding electrode pads that further includesinput/output devices and a power supply. Although only three biosensors125 are depicted in FIG. 1, each of the cavities 111, 113, 115, 117, and119 may have a corresponding biosensor.

In some embodiments, the biosensor 125 is external to the sensing wafer103 and only a patterned sensing surface 139 is provided. The presenceof analyte and/or reaction is detected externally through the one ormore transparent substrates 101 or 103. In such embodiments, the biochip100 may be inserted into a photodetector which would activate thereaction and analyze the extent or identity of the analyte through apositive photodetection at a coordinate on the biochip 100 or a positivephotodetection of a particular type of photon.

As shown in FIG. 1, the patterned sensing surface 139 includes a sensingsurface 141 and a passivation layer 143. Each cavity 111, 113, 115, 117,and 119 may include only sensing surface 141 or passivation layer 143,or both. In some embodiments, the sensing surface 141 or the passivationlayer 143 is the same material as the first surface modification layer145 or the second surface modification layer 147.

FIGS. 2A and 2B are flow charts of various embodiments of methods offabricating a biochip device according to one or more aspects of thepresent disclosure. FIGS. 3-5, 6A-6B, 7-9, 10A-10I, and 11A-11G arecross-sectional views of partially fabricated biochip devicesconstructed according to one or more steps of the method of FIGS. 2A and2B. In operation 201 of FIG. 2A, a fluidic substrate is provided. FIG. 3shows the fluidic substrate 301. The fluidic substrate 301 may be atransparent substrate, including glass or quartz, or opaque, dependingon whether the biochip is designed for external photo-detection orinspection. In some embodiments, the fluidic substrate 301 is silicon,sapphire, silicon carbide, or other commonly used semiconductorsubstrates that does not react with the analyte or solution. Accordingto some embodiments, the fluidic substrate may be at least 100 micronsthick or about 700 microns thick.

In operation 202, a microfluidic channel pattern is formed on thefluidic substrate. FIG. 4 shows a fluidic substrate 301 with amicrofluidic channel pattern. A microfluidic channel pattern is formedfirst by patterning a photoresist over the substrate 301. Afterdeveloping the photoresist and removed portions of the photoresist, theremaining photoresist is used as a etch mask to form the microfluidicchannel pattern in the fluidic substrate 301. The microfluidic channels403, 405, 407, 409, and 411 are etched into the fluidic substrate 301 bywet etch or dry etch. In some embodiments, a dry etch is used to formchannels and wells at about 100 nm deep or deeper. Much deeper channelsand wells that are microns deep or deeper are formed by wet etch or acombination of wet and dry etches. The various channels and wellsconnect to each other in a network of microfluidic channels. Otherfeatures including pumps, valves, sensors, and othermicro-electro-mechanical system (MEMS) devices may be formed or embeddedin the fluidic substrate 301. The field regions between the channels andwells 403, 405, 407, 409, and 411 are island features, for example,features 413, 415, 417, and 419. The fluidic substrate 301 will beeventually bonded to a sensing wafer at top of these island featureswith a bonding material. To avoid leakage between adjacent channels andwells, the bonding material hermetically seals the channels and wellsfrom each other.

In some embodiments, a microfluidic channel pattern is formed on thesensing wafer and not the fluidic substrate. In those embodiments,operation 202 is not performed. In still other embodiments, microfluidicchannel patterns are formed in both the fluidic substrate and thesensing wafer.

In operation 203 of FIG. 2A, a first surface modification layer isdeposited over the microfluidic channel pattern. The first surfacemodification layer 501 conformally covers the microfluidic channelpattern as shown in FIG. 5, including covering the sidewalls of variouswells and channels. The method of depositing the first surfacemodification layer depends on the material. In one example where HMDS isthe first surface modification layer, the HMDS is deposited with aphotolithography tool. In another example where APTES is the firstsurface modification layer, the APTES is deposited by immersing thesubstrate in a solution containing APTES or by exposing the substrate toan APTES vapor. In other examples, the first surface modification layer501 may be deposited using a chemical vapor deposition (CVD) method,including an atomic layer deposition (ALD), plasma enhanced chemicalvapor deposition (PECVD), spraying, coating (including spin-ondeposition), condensing, or printing.

The first surface modification layer may be patterned in optionaloperation 205. In one embodiment, a photoresist pattern is depositedover the fluidic substrate 301 before the first surface modificationlayer is deposited. A patterned first surface modification layer remainsafter the photoresist pattern is removed. In another embodiment, thephotoresist pattern is formed after depositing a blanket first surfacemodification layer. The first surface modification layer is patternedthrough the patterned photoresist by etching, developing, or othermethods of removing. In one example when the first surface modificationlayer is HMDS, a blanket layer is deposited before depositing aphotoresist and patterning the photoresist, then the blanket HMDS isexposed to a solvent that removes the exposed portions of the HMDSthrough the photoresist pattern but not the portions covered by thephotoresist.

In some embodiments, a second surface modification layer pattern isformed on the fluidic substrate using a different method from the firstsurface modification layer. The second surface modification layer may beformed directly on the first surface modification layer or on an exposedportion of the fluidic substrate. For example, a second surfacemodification layer is deposited using a printing method directly over afirst surface modification layer. The second surface modification layermay have a different density and provide a different surface propertyfrom the first surface modification layer. In some embodiments, thesecond surface modification layer may be deposited by selectivedeposition. For example, after patterning the hydrophobic HMDS, ahydrophilic material may be selectively deposited on the exposedsubstrate surface by spraying or coating the substrate as thehydrophilic material would not adhere to a hydrophobic material. FIG. 5shows the second surface modification layer 503 formed over the firstsurface modification layer 501 in some microfluidic wells or channels.

Referring to operation 207 of FIG. 2A, a bonding material pattern isformed on the fluidic substrate. The bonding material pattern is used tobond the sensing wafer and the fluidic substrate. According to variousembodiments, the bonding material is pattern is formed by printingdirectly on the fluidic substrate, by a photolithographic process, or bycondensing a vapor containing the bonding material on the fluidicsubstrate. In some embodiments, the bonding material does not interactwith the first surface modification layer and it is or fully selectivelydeposited directly on the first surface modification layer. In the fulldeposition embodiments, the bonding material may also act as a surfacemodification layer where it is not bonded to the sensing wafer.

FIG. 6A is a cross-sectional diagram of a fluidic substrate 301including a bonding material pattern 601 on portions of the firstsurface modification layer 501. The bonding material acts as barriersbetween microfluidic channels and also seals the biochip at its edges.

In some embodiments, the bonding material can detrimentally react withthe first surface modification layer or the second surface modificationlayer if used. To avoid such a reaction the bonding material is printedonto exposed fluidic substrate portion and the first surfacemodification layer is patterned before the bonding material isdeposited. FIG. 6B is a cross-sectional diagram of a fluidic substrate301 including a bonding material pattern 601 on portions of the fluidicsubstrate 301 without the first surface modification layer 501B. In oneexample where the first surface modification layer is HMDS, the HMDS maybe patterned by photolithography with PR AZ5214-E and developer AZ400K,both of which are commercially available formulations, for example, fromClariant Corporation of Somerville, N.J.

According to various embodiments, the bonding material is asilicon-containing material (for example, a silane) that can beactivated to form silanol or dangling bonds that readily bonds siliconor silicon oxide. In one embodiment, the bonding material is a negativephotoresist polysilsesquioxane (PSQ). PSQ may be patterned by exposing aportion of it to light radiation. The unexposed portion is soluble to aphotoresist developer and washes away during the developing process. PSQis activated using an oxygen-containing plasma. In another embodiment,the bonding material is APTES, which is deposited by vapor and activatedwith a liquid or vapor catalyst, including water, toluene, or phosphatebuffered saline (PBS) solution.

Referring back to FIG. 2A, in operation 209, through-holes are formed inthe fluidic substrate. In some embodiments, the through-holes are formedby laser drilling, microblasting, or ultrasonic drilling. Othertechniques of forming through-holes include various etching techniquesand waterjet drilling. Laser drilling of cylindrical holes generallyoccurs through melting and vaporization of the substrate material, e.g.,ablation, through absorption of energy from a focused laser beam.Depending on the direction of the laser energy, the laser drilledthrough-holes can have an inverse trapezoidal shape in a cross section.Microblasting removes material by driving a high velocity fluid streamof air or inert gases of fine abrasive particles, usually about 0.001inches (0.025 mm) in diameter. Ultrasonic drilling involves using highfrequency vibrations to hammer a bit through materials. At least twothrough-holes are formed for every biochip—an inlet and an outlet. Morethan two through-holes may be used for different inlet fluids or if thebiochip performs separation of the analyte and more than one outlet isused. FIG. 7 is a cross section diagram of a fluid substrate 301 havingtwo through-holes 701 and 703 formed in operation 209.

In operation 211, the bonding material is activated. As discussed, themethod of activation depends on the bonding material. Activation mayinclude exposure to plasma, gas/vapors, fluid, heat, or radiation.Because the fluid substrate is separate from the sensing wafer and doesnot include biological materials, the bonding material activation is notlimited to low energy methods. In the example of PSQ, an oxygen plasmacreates breaks bonds on the PSQ surface and creates dangling bonds whichreadily adhere to a silicon oxide, e.g., silicon dioxide, a thin layerof which is always present on a silicon wafer exposed to ambientconditions. However, some care is taken to not damage the one or moresurface modification layers.

In operation 213, a sensing wafer having two or more modified surfacepatterns having different surface chemistries is provided. The sensingwafer may be formed by the same entity performing method 200, forexample, a semiconductor manufacturer or a semiconductor foundry, or bya different entity, for example, a medical device company or asemiconductor packaging facility. A summary of the sensing waferformation is presented to provide context for the present disclosure. Asensing wafer is usually a silicon substrate with a variety ofbiosensors formed in or on the silicon. One example biosensor is aBioFET, which may have a sensing surface on the front of the gate or onthe back of the gate. A sensing wafer with backside BioFETs have BioFETsconnected by an interconnect structure to electrodes on the back side ofthe sensing wafer. The front side of the sensing wafer would includesensing surfaces on the back side of the gates of the BioFETs. Thesensing surfaces are patterned. The BioFETs are formed directly on thesensing wafer using CMOS fabrication processes. Other biosensors on thesensing wafer may include electrodes, surface plasmon resonance sensor,potentiometric biosensor, and other biosensors that are compatible withconventional CMOS processes. Additional devices may also be formeddirectly on the sensing wafer using conventional CMOS processes, forexample, various MEMS devices may include resonistors, actuators,valves, accelerometers, pressure sensor, heater, cooler, among others.

In operation 215, the sensing wafer and the fluidic substrate is bondedvia the bonding material pattern at a temperature less than about 75degrees Celsius, in some embodiments. The fluidic substrate is alignedwith the sensing wafer and placed together. In some embodiments,pressure is applied to ensure even bonding. In other embodiments, one ormore of the fluidic substrate and the sensing wafer is heated. However,because the surface modification layers and the sensing surface may besensitive to excessive heat, the substrate temperature may be less thanabout 75 degrees Celsius, e.g., about room temperature.

After the fluidic substrate and the sensing wafer are bonded, thecombined workpiece may undergo further processing before the workpieceis diced into individual biochips. Further processing may includetesting, wafer-level packaging, and depositing protective layers. Duringdicing, the workpiece is cut along scribe lines between the biochips. Asufficient width of bonding material is used in the scribe line area toensure that the separated biochips remain sealed.

Some embodiments according to the method 200 of FIG. 2A disclose amicrofluidic channel formed in the fluidic substrate. The microfluidicchannels are formed by dry or wet etching. The microfluidic channels maybe about 100 microns in depth. Because the first surface modificationlayer and the bonding material deposition may be deposited usingsemiconductor tools with different substrate holding mechanisms, whetherthe fluidic substrate has through-holes may limit the application of thefirst surface modification layer or the bonding material. In someembodiments, the first surface modification layer is deposited in ablanket manner over the microfluidic channel and the fluidic substratebefore the jet printing or other selective deposition of the bondingmaterial on the first surface modification layer and then thethrough-holes are formed. In other embodiments, the through-holes areformed before depositing the bonding material, if the jet printer canhandle fluidic substrates having through-holes. In still otherembodiments, the through holes are formed before depositing the firstsurface modification layer. Earlier formation of the through holesensures that less laser drilling by-products remains in the microfluidicchannels.

In certain embodiments, the bonding material is not compatible with thesurface modification layers. The first surface modification layer may bepatterned so that the bonding material deposited does not overlap thesurface modification layer. Whether the surface modification layer ispatterned or not, the through holes may be formed at any stage accordingto the handling capabilities of the tools involved.

In other embodiments, the microfluidic channels are not formed on thefluidic substrate, but rather on the sensing wafer. The surfacemodification layer is deposited in a blanket manner on the fluidicsubstrate and the bonding material may be deposited using aphotolithographic lift-off process before the through holes are formed.In the lift-off process, a photoresist is patterned on the surfacemodification layer creating a pattern of exposed surface modificationlayer. The bonding material is then deposited over the fluidicsubstrate, over both the exposed pattern and the photoresist. Thephotoresist pattern is then removed, or lifted-off, and the portion ofthe bonding material deposited on the exposed portion of the surfacemodification layer remains. In one embodiment where the bonding materialis incompatible or does not adhere to the surface modification layer, anetch may remove the exposed surface modification layer from the fluidicsubstrate before the bonding material is deposited. In anotherembodiment, the through holes may be formed before the photoresistpattern with the bonding material is lifted-off. Of course, bondingmaterial may also be printed using a jet printing just as in otherembodiments.

In some embodiments, washing steps may be used to remove unbounded orexcess bonding material instead of the lift-off process. Bondingmaterial may be selectively bounded by a number of methods includingselective activation through photolithography. Unbounded or excessbonding material may then be washed off the fluidic substrate or thesensing wafer while the bounded bonding material remains.

In still other embodiments, the surface modification layer and thebonding material may be the same material. Examples include PSQ andAPTES. The surface modification layer/bonding material is deposited onthe fluidic substrate and coated with a photoresist or anotherprotective layer for the through-hole formation. Once the through-holesare formed, the protective layer, or the photoresist is removed.

Another aspect of the present disclosure pertains to embodiments wherethe microfluidic channels are formed on the fluidic substrate as opposedto formed in the fluidic substrate. FIGS. 2B and 8-9, 10A-10I, and11A-11G the method 250 and cross-section diagrams in accordance withthis aspect of the present disclosure. Because some of the operations ofmethod 250 are very similar or the same as the operations of method 200,the similarities are not discussed in detail and only differences areemphasized. In operation 251 of method 250 in FIG. 2B, a first substrateis provided. The first substrate may be the fluidic substrate or thesensing wafer with a number of biosensors formed thereon.

In operation 253 a first surface modification layer is patterned on thefirst substrate. The first surface modification layer is deposited onthe first substrate and then patterned using a photolithographicprocess. FIG. 8 shows a first substrate 801 with a first surfacemodification layer 803 deposited thereon. FIG. 9 shows the same firstsubstrate 801 with a patterned first surface modification layer 903 anda patterned photoresist layer 905 over the patterned surfacemodification layer 903. The photoresist is deposited over the firstsurface modification layer 803 and exposed to a light pattern. A portionof the photoresist is then removed. In some embodiments, the firstsurface modification layer may be removed along with the photoresist.For example, HMDS as the first surface modification layer may be removedby developer AZ400K along with photoresist AZ5214-E. In otherembodiments, the first surface modification layer may be etched usingthe photoresist as an etch mask.

In operation 255, a second surface modification layer is patterned onthe first substrate in the first surface modification layer or over thefirst surface modification layer. As shown in FIG. 10A, in someembodiments, a second surface modification layer is deposited over thepatterned photoresist and first surface modification layer of FIG. 9.The second surface modification layer portion 1003 coats the exposedfirst substrate 801 between the patterned photoresist 905 and firstsurface modification layer 903. Another second surface modificationlayer portion 1001 coats the patterned photoresist 905 The patternedphotoresist 905 and the second surface modification layer 1001 over thepatterned photoresist are then removed in a lift-off operation to resultin the cross section of FIG. 10C. FIG. 10C shows the first substrate 801with a layer of first surface modification layer 903 and second surfacemodification layer 1003. In other embodiments, as shown in FIG. 10B, thephotoresist pattern 905 of FIG. 9 is removed to result in simply thefirst substrate 801 with a patterned first surface modification layer903 over it. The second surface modification layer may be selectivelydeposited over the patterned first surface modification layer to resultin the cross section of FIG. 10C when the first surface modificationlayer and the second surface modification layer are insoluble andunwettable with each other. In other words, the two surface modificationlayers have self-assembled surface chemistry. One example of suchcombination is HMDS as one of the surface modification layers and APTESand as the other surface modification layer.

Referring back to method 250 of FIG. 2B, the forming of patternedsurface modification layers may be repeated in loop 271 for additionalsurface modification layers. Additional surface modification layers maybe built over existing surface modification layers or openings may bepatterned and deposited into. Additional surface modification layers mayinclude metal and metal oxide layers, silicon oxide, silicon nitride,and other suitable surface modification layers.

In operation 257, a photoresist is patterned on the first substrate.This operation forms a pattern for the sidewall that lines themicrofluidic channels and wells. The photoresist may be patterned in asimilar process as operation 253, if the openings are formed in thefirst surface modification layer. The photoresist is deposited andexposed to a light pattern. A portion of the photoresist is removedduring the developing process. In operation 259, a portion of the firstsurface modification layer exposed through the patterned photoresist isremoved. In some embodiments, during developing, the developer may alsoremove the underlying first surface modification layer to result incross section of FIG. 10D. FIG. 10D shows the first substrate 801 withpatterned first surface modification layer 903, second surfacemodification layer 1003 and patterned photoresist 1005. In otherembodiments, the first surface modification layer is removed by etchingor other chemical means using the patterned photoresist as a mask toresult in the cross section of FIG. 10D.

Referring back to FIG. 2B, in operation 261 a sidewall structurematerial is deposited over the first substrate. The sidewall structurematerial may be a photoresist, a silicone, a thermoplastic material, orinsulator. The sidewall structure material is deposited sufficientlythick to form the sidewalls of microfluidic channels. According tovarious embodiments, the sidewalls may be 100 microns or taller.Photoresist material can be designed to have different surfaceviscosities and use different deposition process parameters to form arelatively uniform film over the substrate. One such example is SU-8.Silicone material may be molded and shaped to various shapes and sizes.Once hardened, silicone material can provide a hydrophobic orhydrophilic surface as fluid channels. In some embodiments, a siliconematerial polydimethylsiloxane (PDMS) is patterned by a soft lithographyprocess. The PDMS is poured over a wafer having desired sidewall shapesetched into it and hardened. The PDMS is then removed and sealed to aglass substrate by activating the PDMS surface using RF plasma. In otherembodiments, the PDMS is molded using compression molding or injectionmolding directly on the first substrate. Thermoplastic materials canalso be patterned using soft lithography or molding processes. Suitablethermoplastic material includes poly(methyl methacrylate) (PMMA),polycarbonate (PC), and polyimide (PI). Various insulators may bedeposited and patterned using semiconductor processes. For example,silicon oxide, silicon nitride, or various commonly used insulators insemiconductor processing may be used. The sidewall structure materialmay have more than one layers. In some embodiments, the first layer addsthickness while the second or subsequent layers add desirable propertiessuch as adhesion to another substrate and surface property. FIG. 10Eshows a first substrate 801, a patterned first surface modificationlayer 903 and second surface modification layer 1003 over the substrate,a patterned photoresist 1005 over the surface modification layers 903and 1003, and a sidewall structure material 1007 over the firstsubstrate 801 and the patterned photoresist 1005.

Referring back to FIG. 2B, in operation 263 through-holes are formed inthe first substrate. The through-holes are formed using processessimilar to those discussed in association with operation 209 of FIG. 2Awith consideration to the sidewall structure material. Depending on thematerial properties of the sidewall structure material, a differentthrough-hole making process may be used. In one example, if the sidewallstructure material is PMMA and the through-holes are formed using laser,then an annealing step may be required. FIG. 10F shows a cross sectionof the partially fabricated workpiece having through-holes 1009.

Referring back to FIG. 2B, in operation 265 the sidewall structurematerial is patterned. Depending on the material used for the sidewallstructure material, a molded piece or a blanket deposition may be formedon the first substrate in operation 261. In one example, the sidewallstructure material is a photoresist, for example SU-8, which may bepatterned by exposing and developing the photoresist. Other patterningoptions include laser (low power), electron beam, deep UV light, dry orwet etching, and polishing. FIG. 10H shows a cross section of thepartially fabricated workpiece having through-holes 1009, sidewalls 1013and channels 1011. The patterned photoresist is also removed at thisstage or at a later stage, for example, by oxygen plasma after thebonding material is printed.

In some embodiments, the sidewall structure material is patterned beforethe through-hole formation. The patterned photoresist 1005 is used as anetch stop layer. FIG. 10G shows a patterned sidewalls 1013 on a firstsubstrate 801 before the through-holes are formed. From the crosssection of FIG. 10G, the through-holes would then be formed as shown inFIG. 10H.

Referring back to FIG. 2B, in operation 267 a bonding material layer isformed. The bonding material layer may be formed using processes similarto operation 207 of FIG. 2A. FIG. 10I shows a cross section of thepartially fabricated workpiece having bonding material layer on the topof the sidewalls 1013.

Referring back to FIG. 2B, in operation 269 the first substrate tobonded to a second substrate via the bonding material layer. Thisoperation is similar to operation 215 of FIG. 2A. The second substratemay be a sensing wafer if the first substrate is a fluidic substrate.The second substrate may be a fluidic substrate if the first substrateis a sensing wafer. After bonding, a workpiece similar to the biochip100 of FIG. 1 is formed.

As discussed, loop 271 of FIG. 2B may be used to add additional surfacemodification layer on the first substrate. FIGS. 11A to 11G shows suchembodiments. After the first surface modification layer and the secondsurface modification layer are formed, a third surface modificationlayer is deposited by liquid phase coating followed by lithography or UVcuring. In one example, a PEG hydrogel may be UV cured after liquidphase deposition to form the third surface modification layer 1101 ofFIG. 11A. FIG. 11A shows the first substrate 801 with a layer of firstsurface modification layer 903, second surface modification layer 1003in openings of the first surface modification layer 903, and a thirdsurface modification layer 1101 over the second surface modificationlayer 1003. The third surface modification layer may be a self-assembledlayer, i.e., it is selectively deposited if the surface property of thefirst surface modification layer is chosen so that the third surfacemodification layer in liquid phase does not adhere to the first surfacemodification layer. In the example of PEG hydrogel, the hydrogel doesnot adhere to a hydrophobic surface if HDMS is used as the first surfacemodification layer. Various PEG hydrogel may be selected for a itsterminal functional group. For example, diacrylate terminated PEG(PEG-DA) can be used for binding with specific peptides or proteins.

After the third surface modification layer is formed, the processproceeds through operations 257 to 265 as described in association withFIGS. 10D to 10H. FIG. 11B corresponds to FIG. 10D; FIG. 11C correspondsto FIG. 10E; FIG. 11D corresponds to FIG. 10F; FIG. 11E corresponds toFIG. 10G; and FIG. 11F corresponds to FIG. 10H. Referring back to FIG.2B, in operation 267 where a bonding layer is formed on the sidewalls.In the embodiment having a third surface modification layer, the bondingmaterial may also be deposited on the third surface modification layerdepending on the bonding material and method of deposition. For example,if a liquid or vapor phase deposition is used, the liquid or vapor woulddeposit on all surface on which it would adhere. As shown in FIG. 11G,if the first surface modification layer 903 is hydrophobic as thesidewall 1013, then the bonding material 1103 would also deposit ontothe third surface modification layer 1101. In some embodiments, thebonding material may also deposit in the sidewalls of the through-holes.According to various embodiments, for bonding material APTES and thirdsurface modification layer as PEG hydrogel, having a layer APTES doesnot impede biomaterial from entering and functionalizing the PEGhydrogel. As discussed, the photoresist layer over the third surfacemodification layer may be removed after the bonding material depositionin some embodiments. In such case the bonding material would not depositon the third surface modification layer.

One aspect of the present disclosure pertains to a biochip whichincludes a substrate, where the substrate includes at least one holeextending from a first surface of the substrate to a second surface ofthe substrate opposite the first surface, and where the substratecomprises a microfluidic channel pattern. The biochip further includes asurface modification layer over the substrate. Additionally, the biochipincludes a sensing wafer bonded to the substrate, where the sensingwafer has one or more modified surface patterns having different surfaceproperties from each other.

Another aspect of the present disclosure pertains to a biochip includinga fluidic substrate having at least one hole extending therethrough.Additionally, the biochip includes a patterned surface modificationlayer over the fluidic substrate. Moreover, the biochip includes asensing wafer over the fluidic substrate, where the sensing wafercomprises one or more modified surface patterns having different surfaceproperties from each other. Furthermore, the biochip includes a bondingmaterial between the fluidic substrate and the sensing wafer.

In yet another aspect, the present disclosure pertains to biochipincluding a fluidic substrate, where the fluidic substrate has at leastone hole extending from a first surface of the fluidic substrate to asecond surface of the fluidic substrate opposite the first surface.Additionally, the biochip includes a surface modification layer oversidewalls of the at least one hole, wherein the surface modificationlayer includes amino-silane, hexamethyldisilazane (HMDS), agar,hydrogel, or chemistries with thiol functional groups. Moreover, thebiochip includes a sensing wafer over the fluidic substrate.

In describing one or more of these embodiments, the present disclosuremay offer several advantages over prior art devices. In the discussionof the advantages or benefits that follows it should be noted that thesebenefits and/or results may be present is some embodiments, but are notrequired in every embodiment. Further, it is understood that differentembodiments disclosed herein offer different features and advantages,and that various changes, substitutions and alterations may be madewithout departing from the spirit and scope of the present disclosure.

What is claimed is:
 1. A biochip comprising: a substrate, wherein thesubstrate comprises at least one hole extending from a first surface ofthe substrate to a second surface of the substrate opposite the firstsurface and a plurality of cavities, and wherein the substrate comprisesa microfluidic channel pattern configured to connect the at least onehole to each cavity of the plurality of cavities; a plurality of surfacemodification layers over the substrate; and a sensing wafer bonded tothe substrate, wherein the sensing wafer has a plurality of modifiedsurface patterns, wherein a first modified surface pattern of theplurality of modified surface patterns has different surface propertiesfrom a second modified surface pattern of the plurality of modifiedsurface patterns.
 2. The biochip of claim 1, further comprising abonding material between the substrate and the sensing wafer.
 3. Thebiochip of claim 2, wherein the bonding material comprisespoly(phenylmethyl)silsesquioxane (PSQ) or 3-aminopropyl triethoxysilane(APTES).
 4. The biochip of claim 2, wherein the bonding material isconfigured to be non reactive with analyte, bioreceptors, or a testingfluid.
 5. The biochip of claim 1, wherein a first surface modificationlayer of the plurality of surface modification layers comprisesamino-silane, hexamethyldisilazane (HMDS), agar, hydrogel, orchemistries with thiol functional groups.
 6. The biochip of claim 1,wherein at least one surface modification layer of the plurality ofsurface modification layers is over sidewalls of the at least one hole.7. A biochip comprising: a fluidic substrate having at least one holeextending therethrough; a patterned surface modification layer over thefluidic substrate; a sensing wafer over the fluidic substrate, whereinthe sensing wafer comprises a plurality of modified surface patterns,wherein a first modified surface pattern of the plurality of modifiedsurface patterns has different surface properties from a second modifiedsurface pattern of the plurality of modified surface patterns; and abonding material between the fluidic substrate and the sensing wafer. 8.The biochip of claim 7, wherein the bonding material comprisespoly(phenylmethyl)silsesquioxane (PSQ) or 3-aminopropyl triethoxysilane(APTES).
 9. The biochip of claim 7, wherein the fluidic substrate isaligned with the sensing wafer.
 10. The biochip of claim 7, wherein thepatterned surface modification layer comprises amino-silane,hexamethyldisilazane (HMDS), agar, hydrogel, or chemistries with thiolfunctional groups.
 11. The biochip of claim 7, wherein the patternedsurface modification layer is over sidewalls of the at least one hole.12. The biochip of claim 7, wherein the fluidic substrate is configuredto be non reactive with an analyte.
 13. The biochip of claim 7, whereinthe bonding material is configured to be non reactive with an analyte,bioreceptors, or a testing fluid.
 14. A biochip comprising: a fluidicsubstrate, wherein the fluidic substrate has at least one hole extendingfrom a first surface of the fluidic substrate to a second surface of thefluidic substrate opposite the first surface; a surface modificationlayer over sidewalls of the at least one hole, wherein the surfacemodification layer comprises amino-silane, hexamethyldisilazane (HMDS),agar, hydrogel, or chemistries with thiol functional groups; and asensing wafer spaced from the fluidic substrate.
 15. The biochip ofclaim 14, wherein the sensing wafer comprises one or more modifiedsurface patterns, and the modified surface patterns have differentsurface properties from each other.
 16. The biochip of claim 14, furthercomprising a bonding material between the substrate and the sensingwafer.
 17. The biochip of claim 16, wherein the bonding materialcomprises poly(phenylmethyl)silsesquioxane (PSQ) or 3-aminopropyltriethoxysilane (APTES).
 18. The biochip of claim 16, wherein thebonding material is non reactive with an analyte, bioreceptors, or atesting fluid.
 19. The biochip of claim 14, wherein the fluidicsubstrate is aligned with the sensing wafer.
 20. The biochip of claim14, wherein the fluidic substrate has a thickness of at least 100microns (μm).